PROJECT SUMMARY/ABSTRACT New evidence from our studies demonstrates that specific P. acnes ribotypes, distinguished by unique 16S rDNA, may be associated with either healthy skin (ribotype PH) or acne (ribotype PA). We propose to investigate the nature of immune mechanisms by which ribotypes PH and PA induce Th1 vs. Th17-mediated responses. The P. acnes ribotypes PH and PA to be used in this study are the most prevalent and include three each of PH and PA strains that have been enriched in individuals with healthy skin or associated with the acne patient group. We will test our central hypothesis that that P. acnes ribotypes trigger T cells that mediate distinct immune responses, which either contribute to the pathogenesis of acne or lead to a protective mechanism. In Aim 1, we will use informatics, taking advantage of the publicly available skin database DermDB to gain insight into the mechanisms of T cell differentiation. Specifically, using RNAsequencing, we will derive gene signatures of PH and PA-activated innate immune cells, to identify the networks that determine T cell differentiation into functional Th1 and Th17 cell subsets. We will also determine the mechanism(s), including cytokine, by which PH and PA induce T cell differentiation. Although the P. acnes genome has been sequenced, virulence factors and specific ligand(s) that activate immune cells are not clear. We have previously demonstrated that the Toll like receptor-2 (TLR2) mediates the interaction between antigen presenting cells and P. acnes. The bacterial ligand(s) responsible for these interactions are potential therapeutic targets, but are currently unknown. In Aim 2, we propose to use biochemical and proteomic techniques to identify bacterial ligand(s) from PH and PA that lead to differential immune signatures identified in Aim 1, and using siRNA knockdown of specific pathogen recognition receptors (PRR), identify the PRRs that recognize PH and PA to induce the innate cytokines that instruct adaptive immunity. The data generated in Aim 2 will help in the identification of PH and PA ligand(s) that interact with innate PRRs, and provide new insights into the immunobiology of Th1 and Th17 cells. In Aim 3, we will identify the phenotype and function of Th1 vs. Th17 cells induced by P. acnes ribotypes PH and PA. Using the informatics approaches outlined in Aim 1 we have identified IL-26 within the potential new Th17 gene networks. A previously unknown function of Th17- derived IL-26 as a direct antimicrobial agent and activator of DNA-sensing innate immunity was recently reported. Here, we will determine the antimicrobial mechanisms of encoded proteins against P. acnes including IL-26 in in vitro CFU assays, and determine the frequency of IL-26 expressing cells in acne skin specimens by immunohistology. Our investigations of the mechanisms by which distinct ribotypes of a commensal bacterium trigger divergent immune responses will have broad applicability to understanding microbiome-host immune response interactions as well as identify novel targets for immunological intervention. Overall, the K01 training program will allow me to develop the skills and tools needed to become a successful independent investigator.